NADPH- and hydroperoxide-supported 17β-estradiol hydroxylation catalyzed by a variant form (432L, 453S) of human cytochrome P450 1B1

David C. Spink, Barbara C. Spink, Xiaoliang Zhuo, Mirza M. Hussain, John F. Gierthy, Xinxin Ding

Research output: Contribution to journalArticlepeer-review

19 Scopus citations


Human cytochrome P450 1B1 (CYP1B1) catalyzes the hydroxylation of 17β-estradiol (E2) at C-4, with a lesser activity at C-2. The E2 4-hydroxylase activity of human CYP1B1 was first observed in studies of MCF-7 breast cancer cells. Sequencing of polymerase chain reaction products revealed that CYP1B1 expressed in MCF-7 cells was not the previously characterized enzyme but a polymorphic form with leucine substituted for valine at position 432 and serine substituted for asparagine at position 453. To investigate the NADPH- and organic hydroperoxide-supported E2 hydroxylase activities of the 432L, 453S form of human CYP1B1, the MCF-7 CYP1B1 cDNA was cloned and the enzyme was expressed in Sf9 insect cells. In microsomal assays supplemented with human NADPH:cytochrome P450 oxidoreductase, the expressed 432L, 453S form catalyzed NADPH-supported E2 hydroxylation with a similar preference for 4-hydroxylation as the 432V, 453N form, with maximal rates of 1.97 and 0.37 nmol (min)-1(nmol cytochrome P450)-1 for 4- and 2-hydroxylation, respectively. Cumeme hydroperoxide efficiently supported E2 hydroxylation by both the 432V, 453N and 432L, 453S forms at several-fold higher rates than the NADPH-supported activities and with a lesser preference for E2 4- versus 2-hydroxylation (2:1). The hydroperoxide-supported activities of both forms were potently inhibited by the CYP1B1 inhibitor, 3,3',4,4',5,5'-hexachlorobiphenyl. These results indicate that the 432V, 453N and 432L, 453S forms of CYP1B1 have similar catalytic properties for E2 hydroxylation, and that human CYP1B1 is very efficient in catalyzing the hydroperoxide-dependent formation of catecholestrogens. Copyright (C) 2000 Elsevier Science Ltd.

Original languageEnglish (US)
Pages (from-to)11-18
Number of pages8
JournalJournal of Steroid Biochemistry and Molecular Biology
Issue number1-2
StatePublished - 2000


  • Catecholestrogens
  • Cytochrome P450
  • Estrogen metabolism
  • Genetic polymorphism

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Endocrinology
  • Clinical Biochemistry
  • Cell Biology


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